| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1993: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1992: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
DNA fingerprinting requires the use of a specific DNA probe. A universal probe system is a combination of an unlabeled primary probe that has been inserted in a cloning vector and a labeled secondary probe that is specific to the vector. This system is time- and labor-saving in that one does not need to label probes every time one uses them, as long as they are inserted in the same vector.
As first step to a universal probe system, we isolated three DNA fingerprinting probes from a subgenomic library that had been constructed by insertion into the phagemid pUC118 of 1-2 kb fragments from the EcoRI digest of DNA from human myeloma cells (named YA). Sequencing of one of the probes, termed YA1.8 after the length of the insert, revealed that although it contained no apparent repetitive sequence, it detected DNA fingerprints. This suggests that the probe is radically different from the known minisatellite probes ; therefore, we registered it with DDBJ under accession no.D29635.
Next, we isolated single-stranded DNAs of recombinant phagemids, and hybridized them with southern blots of DNAs from father-mother-child trios or unrelated individuals. We were able to detect the DNA hybrids, that is, DNA fingerprints, with a commercially available, alkaline phosphatase-labeled secondary probe or a digoxigenin-labeled universal sequencing primer.
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